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- EMDB-54220: Cryo-EM structure of MATE transporter NorM-VC in complex with dox... -

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Basic information

Entry
Database: EMDB / ID: EMD-54220
TitleCryo-EM structure of MATE transporter NorM-VC in complex with doxorubicin
Map data
Sample
  • Complex: MATE transporter NorM from Vibrio cholerae in complex with NabFab, Nb17_4, anti-Fab nanobody and doxorubicin
    • Protein or peptide: Multidrug resistance protein NorM
    • Protein or peptide: NabFab HC
    • Protein or peptide: NabFab LC
    • Protein or peptide: NorM-Nb17_4
    • Protein or peptide: Anti-Fab nanobody
  • Ligand: DOXORUBICIN
KeywordsNabFab / substrate / multidrug transporter / TRANSPORT PROTEIN
Function / homology: / : / Multi antimicrobial extrusion protein / MatE / antiporter activity / xenobiotic transmembrane transporter activity / monoatomic ion transport / plasma membrane / Multidrug resistance protein NorM
Function and homology information
Biological speciesVibrio cholerae (bacteria) / synthetic construct (others) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsRomane K / Hsieh PY / Kowal J / Locher KP / van Veen HW
Funding support Switzerland, United Kingdom, 2 items
OrganizationGrant numberCountry
Swiss National Science Foundation310030_214834 Switzerland
Biotechnology and Biological Sciences Research Council (BBSRC)BB/K017713/1 United Kingdom
CitationJournal: J Mol Biol / Year: 2025
Title: Doxorubicin recognition and transport by the MATE multidrug transporter NorM from Vibrio cholerae.
Authors: Pei-Yu Hsieh / Ksenija Romane / Julia Kowal / Kaspar P Locher / Hendrik W van Veen /
Abstract: Multidrug and toxic compound extrusion (MATE) transport proteins contribute to multidrug resistance in human pathogens by extruding various cytotoxic compounds from the cellular interior. Despite ...Multidrug and toxic compound extrusion (MATE) transport proteins contribute to multidrug resistance in human pathogens by extruding various cytotoxic compounds from the cellular interior. Despite their importance across all domains of life, the specificities and mechanisms of substrate transport of these proteins remain poorly understood due to limited structural and functional information. Here, we determined the cryo-electron microscopy structure of NorM from Vibrio cholerae (NorM-VC) in complex with the anthracycline antibiotic doxorubicin, using the NabFab approach. The structure reveals that the doxorubicin-binding pocket is located halfway through the membrane, within the C-lobe of the protein. Functional studies targeting the doxorubicin-interacting residues validated the binding pocket and enabled detailed analysis of the doxorubicin transport reaction. Our findings indicate doxorubicin binding within a multisite binding chamber engaged in a general transport mechanism for a variety of substrates.
History
DepositionJul 1, 2025-
Header (metadata) releaseNov 26, 2025-
Map releaseNov 26, 2025-
UpdateNov 26, 2025-
Current statusNov 26, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_54220.png

Downloads & links

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Map

FileDownload / File: emd_54220.map.gz / Format: CCP4 / Size: 343 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.67 Å/pix.
x 448 pix.
= 300.16 Å		0.67 Å/pix.
x 448 pix.
= 300.16 Å		0.67 Å/pix.
x 448 pix.
= 300.16 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.67 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.19
Minimum - Maximum-0.59287035 - 0.9419906
Average (Standard dev.)-0.00020088298 (±0.016759185)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions448448448
Spacing448448448
CellA=B=C: 300.16 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_54220_half_map_1.map
Projections & Slices
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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_54220_half_map_2.map
Projections & Slices
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Sample components

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Entire : MATE transporter NorM from Vibrio cholerae in complex with NabFab...

EntireName: MATE transporter NorM from Vibrio cholerae in complex with NabFab, Nb17_4, anti-Fab nanobody and doxorubicin
Components
  • Complex: MATE transporter NorM from Vibrio cholerae in complex with NabFab, Nb17_4, anti-Fab nanobody and doxorubicin
    • Protein or peptide: Multidrug resistance protein NorM
    • Protein or peptide: NabFab HC
    • Protein or peptide: NabFab LC
    • Protein or peptide: NorM-Nb17_4
    • Protein or peptide: Anti-Fab nanobody
  • Ligand: DOXORUBICIN

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Supramolecule #1: MATE transporter NorM from Vibrio cholerae in complex with NabFab...

SupramoleculeName: MATE transporter NorM from Vibrio cholerae in complex with NabFab, Nb17_4, anti-Fab nanobody and doxorubicin
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5
Source (natural)Organism: Vibrio cholerae (bacteria)
Molecular weightTheoretical: 128 KDa

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Macromolecule #1: Multidrug resistance protein NorM

MacromoleculeName: Multidrug resistance protein NorM / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Vibrio cholerae (bacteria)
Molecular weightTheoretical: 49.892328 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MENSVHRYKK EASNLIKLAT PVLIASVAQT GMGFVDTIMA GGVSAIDMAA VSIAASIWLP SILFGVGLLM ALVPVVAQLN GAGRQHKIP FEVHQGLILA LLVSVPIIAV LFQTQFIIRF MDVEEAMATK TVGYMHAVIF AVPAYLLFQA LRSFTDGMSL T KPAMVIGF ...String:
MENSVHRYKK EASNLIKLAT PVLIASVAQT GMGFVDTIMA GGVSAIDMAA VSIAASIWLP SILFGVGLLM ALVPVVAQLN GAGRQHKIP FEVHQGLILA LLVSVPIIAV LFQTQFIIRF MDVEEAMATK TVGYMHAVIF AVPAYLLFQA LRSFTDGMSL T KPAMVIGF IGLLLNIPLN WIFVYGKFGA PELGGVGCGV ATAIVYWIML LLLLFYIVTS KRLAHVKVFE TFHKPQPKEL IR LFRLGFP VAAALFFEVT LFAVVALLVA PLGSTVVAAH QVALNFSSLV FMFPMSIGAA VSIRVGHKLG EQDTKGAAIA ANV GLMTGL ATACITALLT VLFREQIALL YTENQVVVAL AMQLLLFAAI YQCMDAVQVV AAGSLRGYKD MTAIFHRTFI SYWV LGLPT GYILGMTNWL TEQPLGAKGF WLGFIIGLSA AALMLGQRLY WLQKQSDDVQ LHLAAK

UniProtKB: Multidrug resistance protein NorM

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Macromolecule #2: NabFab HC

MacromoleculeName: NabFab HC / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 25.684463 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: EISEVQLVES GGGLVQPGGS LRLSCAASGF NFSYYSIHWV RQAPGKGLEW VAYISSSSSY TSYADSVKGR FTISADTSKN TAYLQMNSL RAEDTAVYYC ARGYQYWQYH ASWYWNGGLD YWGQGTLVTV SSASTKGPSV FPLAPSSKST SGGTAALGCL V KDYFPEPV ...String:
EISEVQLVES GGGLVQPGGS LRLSCAASGF NFSYYSIHWV RQAPGKGLEW VAYISSSSSY TSYADSVKGR FTISADTSKN TAYLQMNSL RAEDTAVYYC ARGYQYWQYH ASWYWNGGLD YWGQGTLVTV SSASTKGPSV FPLAPSSKST SGGTAALGCL V KDYFPEPV TVSWNSGALT SGVHTFPAVL QSSGLYSLSS VVTVPSSSLG TQTYICNVNH KPSNTKVDKK VEPKSCDKTH T

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Macromolecule #3: NabFab LC

MacromoleculeName: NabFab LC / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 23.258783 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SDIQMTQSPS SLSASVGDRV TITCRASQSV SSAVAWYQQK PGKAPKLLIY SASSLYSGVP SRFSGSRSGT DFTLTISSLQ PEDFATYYC QQSSSSLITF GQGTKVEIKR TVAAPSVFIF PPSDSQLKSG TASVVCLLNN FYPREAKVQW KVDNALQSGN S QESVTEQD ...String:
SDIQMTQSPS SLSASVGDRV TITCRASQSV SSAVAWYQQK PGKAPKLLIY SASSLYSGVP SRFSGSRSGT DFTLTISSLQ PEDFATYYC QQSSSSLITF GQGTKVEIKR TVAAPSVFIF PPSDSQLKSG TASVVCLLNN FYPREAKVQW KVDNALQSGN S QESVTEQD SKDSTYSLSS TLTLSKADYE KHKVYACEVT HQGLSSPVTK SFNRGEC

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Macromolecule #4: NorM-Nb17_4

MacromoleculeName: NorM-Nb17_4 / type: protein_or_peptide / ID: 4 / Details: Nanobody / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 14.356979 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
QRQLVESGGG LVQPGGSLRL SCAASGIIFK INDMGWFRQA PGKEREGVAG ITSGGRTNYA DSVKGRFIIS RDNVKNTVYL QMNSLEPED TAVYYCKSDG LISYAASQLS TYWGKGTPVT VSSHHHHHHE PEA

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Macromolecule #5: Anti-Fab nanobody

MacromoleculeName: Anti-Fab nanobody / type: protein_or_peptide / ID: 5 / Details: Nanobody / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 13.390644 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
GSQVQLQESG GGLVQPGGSL RLSCAASGRT ISRYAMSWFR QAPGKEREFV AVARRSGDGA FYADSVQGRF TVSRDDAKNT VYLQMNSLK PEDTAVYYCA IDSDTFYSGS YDYWGQGTQV TVSS

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Macromolecule #6: DOXORUBICIN

MacromoleculeName: DOXORUBICIN / type: ligand / ID: 6 / Number of copies: 1 / Formula: DM2
Molecular weightTheoretical: 543.519 Da
Chemical component information

ChemComp-DM2:
DOXORUBICIN

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 %

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 65.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.6 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 327809
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

SoftwareName: Coot
Output model

PDB-9rsj:
Cryo-EM structure of MATE transporter NorM-VC in complex with doxorubicin

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